Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide. These tumors have aggressive phenotypes and high recurrence, generally caused by the growth of multiple individual neoplasms of different clonalities. This pattern of growth and recurrence suggests that the oncogenic events involve a large proportion of hepatocytes. Recent data from experimental models and humans indicate that in preneoplastic livers apoptosis is associated with hepatocyte replication and that tumor promoters act at this stage by decreasing apoptosis rather than changing the rate of cell proliferation. Nevertheless, the precise role of apoptosis in tumorigenesis remains unclear. The investigator has studied in detail the development of HCC in transgenic mice that overexpress Transforming Growth Factor alpha (TGFa). In this model approximately 80% of the mice develop HCC after 12 months of age. Tumor development is preceded by an initial phase of hepatocyte proliferation and organ growth followed by a period in which both proliferation and apoptosis are high but organ size is normal. Shortly after that, diploid hepatocytes emerge and eventually produce tumors. Work by the investigator in his laboratory and in collaborative efforts have shown that blockage of NF-kB in hepatocyte cell lines obtained from TGFa transgenic mice causes massive apoptosis and that tumors appear to originate from a population of diploid hepatocytes containing aneuploid cells which might be less sensitive to apoptosis. The investigator postulates that during liver carcinogenesis, proliferation and apoptosis may be interconnected through pathways that involve NF-kB activation and that blockage of one of these pathways leads to cell death. The specific aims of the proposal are to determine: 1) whether blockage of NF-kB causes apoptosis in nontumorigenic and tumorigenic replicating hepatocytes; 2) whether diploid hepatocyte populations that emerge at a stage of liver carcinogenesis prior to tumor development are resistant to apoptosis; 3) whether blockage of apoptosis during liver carcinogenesis causes organ enlargement and accelerated tumorigenesis. Unique differentiated tumorigenic and nontumorigenic hepatocyte cell lines developed in the investigator s laboratory from TGFa transgenic mice as well as p53 null mice will be transfected with IkBa and TRAF2 deletion mutants that act as dominant negative regulators of NF-kB activation to determine whether apoptosis may result from NF-kB blockage. The relationships between cell proliferation and apoptosis during hepatocarcinogenesis in vivo will be studied in TGFa transgenic mice and in TGFa/Bcl-2 double transgenics in which hepatocytes express both an endogenous mitogen and an antiapoptotic gene. It is expected that the results from these experiments will lead to a better understanding of the pathogenesis of HCC and will have important implications for devising new therapeutic strategies for these tumors.